Principal Investigator/Program Director (Last, First, Middle: Chen, Ridong Abstract Due to its narrow time-window of administration (up to 4.5 hours post symptoms) and 6-7 fold increased risk of intracranial hemorrhage, merely 3-5% of acute ischemic stropke patients receive recombinant tissue plasminogen activator (tPA), the only FDA-approved drug for this indication. When tPA is given beyond 4.5 hours of stroke onset, deleterious effects of the drug ensue, especially, hemorrhagic transformation (HT), which causes the most significant morbidity and mortality in stroke patients. In animal models, treatment with IL-2 induced regulatory T cells that improved stroke outcomes while attenuating HT. Recent clinical trials have demonstrated that low-dose (1-3% of the cancer dose) IL-2 therapy was well tolerated and improved clinical outcomes in patients with autoimmune or inflammatory diseases without causing immunosuppression or increasing infection. Clearly, treatment with low-dose IL-2 to induce regulatory T cells carries a high therapeutic potential to improve the efficacy, safety and expand time window of tPA for stroke patients. Current IL2 therapy (Aldesleukin, 15kD) is an unglycosylated recombinant IL2 from E coli. Rapid renal elimination results in a very short in vivo half-life of IL-2 in the range of minutes due to its small molecular size. This has been a major limitation for IL-2-based strategies, thus requiring the use of high and multiple doses of IL-2 and causing toxic side effects. We have designed and produced a novel IL2-based therapy. In this study, we will follow the STAIR recommendations and RIGOR guidelines to conduct the Phase I preclinical validation of tPA and the proprietary IL2 analog in male and female animals as innovative combination therapy for stroke. Specific Aim. Determine whether combining tPA with the proprietary IL2 analog will improve long-term outcomes compared to the vehicle and tPA in the thrombo-embolic stroke model of rats 6h after occlusion.